Use of essential oil feed additives in ruminant nutrition: Opportunities and challenges

Datum: 2016-12-19

Remarks by Chaouki Benchaar, Agriculture and Agri-Food Canada, Canada, during the ruminant breakout session at the 2016 World Nutrition Forum in Vancouver, Canada.

Introduction

In livestock production, antibiotics at sub-therapeutic levels are commonly used to enhance the efficiency of converting feeds to gain (e.g., milk and meat) and/or prevent metabolic disorders and health problems. However, the restriction and the ban (e.g., European Union) on the use of antibiotics in several countries have prompted scientists and the feed industry to search for alternative products. Phytogenics offer a unique opportunity in this regard as many plants produce secondary metabolites, such as essential oils that, when extracted and concentrated, may exert antimicrobial activities against a wide variety of rumen microorganisms. Accordingly, research on the use of essential oils in ruminant nutrition has increased over the last decade. Understandably, most of this research is laboratory based (i.e., in vitro) and has allowed to screen a large number of essential oils and their main constituents for their effects on rumen microbial fermentation. The number of in vivo studies has increased over the last years, but more research work is still required to assess the potential of these phytogenics to enhance feed efficiency (nitrogen and energy) and improve ruminant performance (i.e., milk and meat). This review presents recent developments in use of essential oils as feed additives in ruminant nutrition. Antimicrobial properties, mechanisms of action, effects on ruminal protein metabolism, enteric methane production, animal performance, and challenges are discussed.

Essential oils: Definition, chemistry, and antimicrobial properties

Essential oils (also referred as volatile or ether oils) are complex mixtures of volatile lipophilic secondary metabolites. They are typically extracted from plant material by boiling water and steam distillation, but other methods also include solvent extraction, supercritical CO2 extraction, and expression extraction (Benchaar and Greathead, 2011). Essential oils are plant specific and are responsible for a plant’s characteristic flavor and fragrance (Greathead, 2003). They also play an important role in protecting the plant from abiotic and biotic stressors and acting as attractants to organisms that pollinate and disperse seeds (Wink and Schimmer, 1999). Essential oils can be extracted from all parts of a plant including leaves, roots, flowers, petals, seeds, fruits, stems and barks. The yield and the composition of essential oils can vary widely among plants depending on plant species (Martínez et al., 2006), the geographical location (e.g., climatic conditions, soils; Vokou et al., 1993), part of the plant (e.g., seeds vs. leaves; Delaquis et al., 2002), plant health (e.g., insect attacks; Chen, 2008), and the method of extraction (Anitescu et al., 1997).

Chemically, essential oils are typically composed of phenylpropene and terpene secondary metabolites. The metabolic pathways of biosynthesis of these 2 groups are presented in Figure 1. The phenylpropenes are synthesized via the shikimate pathway (Sangwan et al., 2001), while the terpenes (monoterpenes and sesquiterpenes) are synthesized using the citric acid cycle intermediate acetyl-CoA (mevalonate pathway) and the glycolytic intermediates glyceraldehydes 3-phosphate and pyruvate (deoxyxylulose pathway) as precursors (Dewick, 2002).

Essential oils have been shown to exhibit antimicrobial activity (Cowan, 1999, Burt, 2004). Because of the large number of components present in essential oils, it is most likely that their antibacterial activity is not due to a single mode of action but involves several targets in the cell (Burt, 2004, Acamovic and Brooker, 2005). The antibacterial activity of essential oils is mainly related to their hydrophobicity, which enables them to accumulate in the lipid bilayer of the bacterial plasma membrane from where they exert their effects, which vary according to type of secondary metabolite. Some alter membrane permeability, some interact with membrane proteins, and others may interact directly with cytoplasmic components from within the plasma membrane or by diffusing into the cytoplasm itself (Figure 2). Not all of these mechanisms are separate targets; some are affected as a consequence of another mechanism being targeted (Burt, 2004).

Gram-positive bacteria appear to be more susceptible than Gram-negative bacteria to the antibacterial properties of essential oils (Burt, 2004). This may be expected as Gram-negative bacteria have an outer layer surrounding their cell wall that acts as a permeability barrier, limiting the access of hydrophobic compounds (Cosentino et al., 1999, Lambert et al., 2001, Burt, 2004). However, Helander et al. (1998) reported that the phenolics thymol and caravacrol also inhibited growth of Gram-negative bacteria by disrupting the outer cell membrane. It appears that the small molecular weight of essential oils allows them to pass through porin proteins in the outer membrane and gain access to the plasma membrane (Nikaido, 1994, Dorman and Deans, 2000).

Effects of essential oils on rumen microbial fermentation

Because essential oils are relatively novel feed additives in ruminant nutrition, their effects on ruminal fermentation and overall nutritional effects remain not well defined. The range of essential oils and their components available is extensive (more than 3000; Van de Braak and Leijten, 1999). As a consequence, researchers have heavily relied on short-term laboratory studies (i.e., in vitro) to evaluate their potential to manipulate ruminal fermentation and predict in vivo effects. Accordingly, several in vitro studies used batch or continuous culture systems to screen the large number of essential oils with the ultimate objective to identify compounds showing promises for in vivo application.

Effects on ruminal protein metabolism

Ruminant animals are relatively inefficient users of dietary nitrogen (N). The efficiency of transfer of feed N into milk protein varies between 25 to 30% (Hristov et al., 2005, Lapierre et al., 2005), as the remaining N is being excreted in the environment with urine and feces. Consequently, N not used for milk protein synthesis or gain contributes to pollution of air and ground and surface water. Thus, improving the efficiency of nitrogen utilization in ruminants has a positive effect on animal production and the environment.

Several in vitro studies have been conducted to determine the effects of essential oils and their main constituents on rumen protein metabolism. Results from studies are variable depending on the essential oil (or the essential oil compound), the dose and the in vitro technique (batch versus continuous culture) used. For instance, Castillejos et al. (2006, 2008) conducted a series of batch culture and continuous culture experiments to assess the effects of thymol on rumen microbial fermentation. At doses of 5 and 50 mg/l, thymol had no effect on ruminal N metabolism. At higher doses (500 and 5000 mg/l), thymol decreased ammonia concentration in 24-h batch cultures, which was consistent with the inhibition of the deamination process observed in the same study. However, when added at 500 mg/l in a continuous culture system, thymol increased the concentration of large peptide N and small peptide plus amino acid N, but had no effect on ammonia concentration. These changes are an indication that both proteolysis and peptidolysis processes were stimulated by thymol. A number of studies have investigated in vivo the effects of essential oils on ruminal N metabolism. Chaves et al. (2008b) observed no change in ruminal ammonia concentration in lambs supplemented with cinnamaldehyde or carvacrol (200 mg/kg of dry matter intake [DMI]). Likewise, Yang et al. (2010a, 2010b) observed no change in ruminal ammonia concentration in feedlot cattle fed 400, 800 or 1600 mg/d of either eugenol or cinnamaldehyde. Feeding cinnamon oil (50 mg/kg DMI), cinnamaldehyde (43 and 50 mg/kg DMI), or eugenol (25, 50 and 75 mg/kg DMI) to dairy cows had no effect on ruminal protein degradation and ammonia concentration (Benchaar et al., 2008b, 2012, 2015, Benchaar, 2016). Tekippe et al. (2011) reported that pulse-dosing 500 g/day of oregano leaves (90.8% carvacrol of total essential oil) in the rumen of dairy cows increased ruminal ammonia concentration. In contrast, Hristov et al. (2013) reported a decrease in ruminal ammonia concentration in cows fed 250, 500 or 750 g/day of oregano leaves (78.5% carvacrol of total essential oil).

Effects on enteric methane production

Methane (CH4) is a potent greenhouse gas produced primarily in the rumen during microbial feed digestion (i.e., enteric fermentation) and to a lesser extent from manure storage. According to Gerber et al. (2013), livestock contributes 44% of the global anthropogenic CH4 emissions with most of it from enteric fermentation by ruminants. Enteric CH4 is also a significant loss (2 to 12% of the gross energy ingested) in productive energy to the ruminant (Johnson and Johnson, 1995). Therefore, the interest in reducing enteric CH4 production in the rumen is well-justified from nutritional and environmental standpoints.

A number of in vitro studies have evaluated the potential of essential oils to inhibit ruminal methanogenesis. The reported effects varied with the type and the dose of the essential oil used. Macheboeuf et al. (2008) assessed in 16-h batch culture incubations the effects of thyme (470 g/kg thymol, 200 g/kg terpinene and 200 g/kg p-cymene) on ruminal fermentation. A minimum of 300 mg/l of thymol provided as is, or via thyme oil, was required to inhibit CH4 production with a concomitant decrease in total volatile fatty acid (VFA), acetate and propionate production, a reflection of an inhibition of feed degradation. The extent of CH4 reduction was more pronounced with thyme oil (providing 300 mg/l of thymol) as when thymol was provided alone at the same concentration (-62 versus -32%, respectively), which suggests that other constituents in thyme oil contributed to its antimicrobial activity.

Castro-Montoya et al. (2012) observed a strong decrease (90 to 95%) in CH4 production when cinnamaldehyde was added at 1.5 and 3 mg/ml in 24-h batch culture incubations. This reduction occurred with a concomitant decrease in total VFA production, an indication of an inhibition of feed degradation. In a previous in vitro study by Pellikaan et al. (2011), CH4 production was almost completely inhibited during the first 30 h of 72 h incubation when cinnamaldehyde (2.5 mg/ml) was added to soybean hulls or corn. Interesting, the extent of decrease of CH4 production from soybean hulls was only 65% after 72 h incubation, suggesting a possible adaptation of rumen microbes to cinnamaldehyde exposure.

Busquet et al. (2005a) evaluated in vitro the effects of garlic oil and 2 of its compounds, diallyl disulphide and allyl mercaptan, on CH4 production. At 300 mg/l, allyl mercaptan decreased CH4 production by 19.5%, and total VFA concentration, without altering digestibility. At the same concentration, garlic and diallyl disulphide reduced CH4 production by -74 and -69% respectively, but dry matter digestibility and total VFA concentration were also depressed. That diallyl disulphide reduced CH4 production to the same extent as the oil fraction of garlic may indicate that this sulphur containing compound is responsible for most of the antimethanogenic activity of garlic oil.

A limited number of studies have examined in vivo the effects of essential oils and their main components on enteric CH4 production by ruminants. Beauchemin and McGinn (2006) observed no change in CH4 production, although feed digestibility decreased in beef cattle supplemented with 1 g/day of a commercial mixture of essential compounds (Crina® Ruminant, Akzo Nobel Ltd.). McIntosh et al. (2003) observed that the inhibition of the growth of the methanogen Methanobrevibacter smithii occurred only when the concentration of Crina® Ruminant exceeded 1000 mg/l. This level is 33-times higher than that used by Beauchemin and McGinn (2006), a feeding rate that is not practical, due to potentially adverse effects on efficiency of ruminal fermentation and diet digestibility. Tekippe et al. (2011) and Hristov et al. (2013) observed a reduction in enteric CH4 production of dairy cows fed or ruminally pulse-dosed with oregano leaves. Meale et al. (2014) observed no change in CH4 production of cows fed garlic (5 g/day) or juniper berry (2 g/day) oils. More recently, Benchaar (2016) observed no effects of feeding (50 mg/kg DMI) cinnamon oil or cinnamaldehyde to dairy cows on enteric CH4 production.

Effects on animal performance

Compared to the large number of in vitro studies published to date on the effects of essentials oils on rumen microbial fermentation, the number of in vivo studies that have examined the effects of these phytogenics on ruminant performance (milk and gain) remains relatively low.

A series of studies by Benchaar et al. (2008b, 2012, 2015) and Benchaar (2016) revealed no effect of feeding cinnamon oil (50 mg/day), cinnamaldehyde (50 mg/kg DMI) or eugenol (25, 50 and 75 mg/kg DMI) to dairy cows on milk yield and milk composition. Yang et al. (2007) observed that feeding garlic (5 g/day) and juniper berry (2 g/day) oils to dairy cows had no effect on milk production and milk composition. Tager and Krause (2011) found no changes in milk performance of dairy cows fed 0.5 or 10 g/day of a mixture of cinnamaldehyde and eugenol. Hristov et al. (2013) observed no changes in milk production of dairy cows fed oregano leaves despite an increase in DMI.

Based on results of studies cited above, it appears that the effects of essential oils and their main components on ruminant performance were not conclusive, which is not surprising considering the equivocal effects of these secondary metabolites on DMI and ruminal fermentation characteristics. In some studies, the improved performance of animals supplemented with essential oils has been related to higher feed intakes rather than to an alteration of rumen microbial fermentation and nutrient digestibility.

Challenges

Most of the research on essential oils published to date is of short term nature and has been conducted using in vitro systems (Calsamiglia et al., 2007, Benchaar et al., 2008a, Benchaar et al., 2009). Although the number of in vivo studies has increased over the last years, more research work is still needed to determine the potential of essential oils to improve feed (N and energy) efficiency and improve animal performance.

Based on results published to date, it appears that phenolic compounds (i.e., thymol, eugenol, carvacrol) or essential oils with high concentrations of these phenolics, cinnamon oil and its main component cinnamaldehyde, garlic essential oil and its derivatives, in particular diallyl disulphide, and other essential oils may be effective, at least in vitro, in improving efficiency on N and energy utilization in ruminants.

However, the use of essential oils in ruminant nutrition could be limited for following reasons:

From the published in vitro studies, it appears that high concentrations of essential oils (i.e., >300 mg/L of culture fluid) are required to alter ruminal fermentation. These levels are too high to be achieved in vivo and are impractical in terms of feeding due to potentially deleterious effects on palatability and efficiency of ruminal fermentation (Benchaar et al., 2008a, Benchaar et al., 2009).

Because of the wide spectrum of activity of essential oils and because of the high dosage required to alter ruminal fermentation, reported beneficial effects on ruminal fermentation (i.e., N metabolism and CH4 energy losses) were often offset by an overall inhibition of fermentation and feed digestion which, of course, is not desirable due to negative consequences on animal productivity.

Rumen microbes have been shown to have the capacity to adapt to and/or degrade a wide variety of plant secondary metabolites such as saponins and tannins (Newbold et al., 1997, Makkar, 2003) and the same appears to be true for essential oils as shown in some studies (McIntosh et al., 2003, Busquet et al., 2005b). This adaptive response may explain why, to date, responses in vitro (i.e., short term exposure) are not as marked as in vivo (i.e., long term exposure). The adaptation of rumen microbes to essential oils represents a major challenge for use of these phytogenics in ruminant nutrition.

Despite the increased interest in using essential oils to manipulate ruminal fermentation, little research has been completed to determine the fate of essential oils and their compounds in the gastro-intestinal tract of ruminants (Benchaar and Greathead, 2011). There are indications that some essential oils, such as terpenes, can be degraded within the gastro-intestinal tract. Cluff et al. (1982) reported that 80% of the monoterpenoids contained in sagebrush disappeared from the rumen of wild mule deer. Likewise, Malecky et al. (2009) reported high rates of disappearance (80 to 100%) of the terpenes linalool, p-cymene and α- and β-pinene from the rumen of dairy goats. Different mechanisms have been proposed to explain the disappearance of essential oil terpenes from the digestive tract. These include, bioconversion by the rumen microflora (Welch and Pederson, 1981, Schlichtherle-Cerny et al., 2004), transfer to the gas phase of the rumen due to volatility of terpenes resulting in their loss during eructation (Cluff et al., 1982, White et al., 1982), and absorption across the ruminal wall into the blood system and excretion in the urine (Michiels et al., 2008, Malecky et al., 2009). The degradation and/or conversion of terpenes within the gastrointestinal tract may explain the discrepancy between in vitro and in vivo studies (Benchaar et al., 2008b and 2009).

Although several essential oils are considered safe, some research has shown that these compounds can be toxic. For example, sulphur-containing compounds of garlic and onion have been shown to exert hemotoxic effects in beef cattle (Rae, 1999) and horses (Pearson et al., 2005). Use of essential oils as feed additives in livestock production must also be safe for the feed manufacturing personnel and farm workers. Indeed, these compounds have been reported to be potentially irritating and thus, causing allergic dermatitis (Burt, 2004), suggesting that caution should be taken by users in handling of such feed additives.

Conclusions

The well documented antimicrobial activity of essential oils has prompted researchers to examine the potential of these phytogenics to favorably modify rumen microbial populations in order to enhance efficiency of ruminal fermentation and to improve nutrient utilisation in ruminants. Results from laboratory-based studies (i.e., in vitro) revealed the potential of phenolic compounds (i.e., thymol, eugenol, carvacrol) or essential oils with high concentrations of phenolics, cinnamon oil and its main component (i.e., cinnamaldehyde), garlic and its derivatives (e.g., diallyl disulphide) to improve N and/or energy utilization in ruminants. However, results from in vivo studies were less conclusive because of the different dosage applied compared to in vitro studies. The capacity of rumen microbes to adapt to and degrade essential oils represents one of the main challenges for the use of essential oil additives in ruminant nutrition.